Species-specific restriction factors have attracted exceptional research interest from HIV/AIDS researchers. This kind of innate immunity is mediated by proteins such as APOBEC3 proteins, TRIM5alpha, TRIMCyp, and BST2/Tetherin. To infect new species, retroviruses must evolve specific countermeasures. Remarkably, it is now believed that the accessory genes (vif, vpx, vpu, nef, and probably vpr) of primate lentiviruses are largely devoted to countering the antiviral effects of species-specific restrictio factors. There is intensive interest in therapeutic application of restriction factor science via gne therapy or small molecule approaches. Much remains unknown about how these factors actually function in vivo, in a body. This includes whether they can be exploited to protect against trans-mucosal or percutaneous infection. The idea is compelling because these factors can act immediately, before signaling cascades and/or adaptive immunity are mounted, and in a cell-autonomous way. The experiments in this field have largely been conducted in cultured cells of diverse species, with great success. There are also limited observations of allelic influences in macaques and humans (and, previously, the Fv1 system for mouse retroviruses). However, controlled body-level genetic manipulation of a susceptible species has not been possible for lentiviruses. We do not know how much a single restriction factor or a combination of them can defend a host and at which of three broadly considered levels: initial infection, development of systemic viremia, and disease (AIDS). Restriction factors have never been experimentally tested by introduction into the germline of a species susceptible to a lentivirus. Here we will carry out such experiments for the first time. We will do this in a natural lentiviral host that develops AIDS and is afflicted with a pandemic AIDS virus. We enabled efficient cat transgenesis to make this possible. We include the results of this pilot project as preliminary data for this R01 proposal. Unlike rodents, the domestic cat is naturally susceptible to an AIDS virus. Moreover, FIV is restricted by primate restriction factors, including both Old and New World TRIMCyps. This is an ideal use of this AIDS model. It will enable experiments that cannot be done in mice, rats, rabbits, macaques or people. We will determine the ability of restriction factor(s) to block FIV at the three potential levels described above. We will answer whether a species can be rendered genetically immune to its AIDS virus by addition of restriction factor genes. We will also determine whether and how the virus evolves in response. We will determine, in turn, if protected cell lineages are selected and enriched in vivo as the virus replicates. Thus, this project is innovative at the basic science level and the translational level We believe we are an ideal lab to do this because we have years of experience working with the cat and analyzing its tissues, we have specific experience with FIV and HIV, we have background and knowledge in host cell factors and restriction, and we enabled efficient feline transgenesis.